An amperometric biosensor for determination of biochemical oxygen demand in wastewater has been developed to overcome the time consuming monitoring procedures. The performance and stability of the immobilized membrane have been investigated at 37°C and pH 6.8. Immobilized microbial membranes maintain their stability and activity after intermittent use for 400 cycles when stored at 4°C in sodium phosphate buffer pH 6.8. The response time of the BOD sensor was only 90min, being independent of the concentration, and the lower detection limit was 1mg/l. The obtained BOD values showed correlation with that of the conventional method for BOD determination (BOD5) with a deviation of ±10%. Moreover, the sensor exhibits good repeatability (3.39–4.45%) and reproducibility (1.85–2.25%). Software was added to upgrade this sensor and to make it a promising candidate for online monitoring.